Haplotype partitioning in the proximal promoter of the human growth hormone (GH1) gene

Abstract

The invention relates to variants of the human growth gene (GH1) and, in particular, variants in the proximal promoter region thereof. Moreover, the invention relates to the interaction of said variants and how said interaction affects growth hormone expression.

We claim:  
     
         1 . A method for diagnosing the existence of, or a susceptibility to, growth hormone dysfunction in an individual comprising: 
 (a) obtaining a test sample of a nucleic acid molecule encoding the proximal promoter region of the growth hormone gene (GH1) from an individual to be tested;    (b) examining said nucleic acid molecule for a plurality of the following six SNP's: 1, 6, 7, 9, 11 and 14 (described in Table 1), or the corresponding haplotypes thereof (also described in Table 1); or a polymorphism in linkage disequilibrium therewith;    (c) and where a plurality of said SNP's, or their said corresponding haplotypes, or their said corresponding polymorphisms, exist determining that the individual may be suffering from, or has a susceptibility to, growth hormone dysfunction.    
     
     
         2 . A method according to  claim 1  wherein said polymorphism is at 114 of the locus control region of the said gene.  
     
     
         3 . A method according to  claim 1  wherein said polymorphism is at 1194 of the locus control region of said gene.  
     
     
         4 . A method for diagnosing the existence of, or a susceptibility to, growth hormone dysfunction in an individual, comprising: 
 (a) obtaining a test sample of a nucleic acid molecule encoding the proximal promoter region of the growth hormone gene (GH1) from an individual to be tested;    (b) examining said nucleic acid molecule for any one or more of the following haplotypes in Table 1 indicated as numbers 3, 4, 5, 7, 11, 13, 17, 19, 23, 24, 26 or 29;    (c) and where said haplotypes exist determine that the individual may be suffering from, or has a susceptibility to, growth hormone dysfunction.    
     
     
         5 . A method according to  claim 1  wherein said examining step under (b) above comprises PCR amplification of said gene.  
     
     
         6 . A method according to  claim 5  wherein the primer is selected from the group consisting of:  
       
         
           
                 
                 
               
                     
                     
                 
                     
                   GGG AGC CCC AGC AAT GC (GH1F);  
                 
                     
                   and 
                 
                     
                     
                 
                     
                   TGT AGG AAG TCT GGG GTG C (GH1R). 
                 
                     
                     
                 
             
                
                
                
                
                
                
               
            
           
         
       
     
     
         7 . A method according to  claim 6  wherein said primers are labelled in order to facilitate detection of the amplified product.  
     
     
         8 . A kit suitable for carrying out diagnostic methods of  claim 1 , which kit comprises: 
 (a) at least one of the following primers for detecting and/or amplifying the proximal promoter region of the growth hormone gene (GH1);                                      GGG AGC CCC AGC AAT GC (GH1F);                       TGT AGG AAG TCT GGG GTG C (GH1R);                                 and, optionally,    (b) one or more reagents suitable for carrying out PCR for amplifying desired regions of the patient's DNA.    
     
     
         9 . A kit according to  claim 8  wherein additional primers are used that are complementary to selected regions of the gene containing the SNP's defined herein as 1, 6, 7, 9, 11 and 14.  
     
     
         10 . A vector comprising at least the proximal promoter region of GH1 wherein said region comprises a plurality of the following SNP's: 1, 6, 7, 9, 11 and 14.  
     
     
         11 . A vector according to  claim 10  wherein said region comprises SNP's 6 and 9.  
     
     
         12 . A vector comprising at least the proximal promoter region of GH1 wherein said region comprises SNP's 10 and 12.  
     
     
         13 . A vector comprising at least the proximal promoter region of GH1 wherein said region comprises SNP's 8 and 11.  
     
     
         14 . A vector according to  claim 10  wherein said region is characterised by any one or more of the following haplotypes shown in Table 1: 3, 4, 5, 7, 11, 13, 17, 19, 23, 24, 26 or 29.  
     
     
         15 . A vector according to  claim 10  which further comprises a GH1 locus control region proximal promoter fusion construct.  
     
     
         16 . A vector according to  claim 10  wherein said proximal promoter region is functionally linked to the coding region of a selected gene wherein the activity of the said proximal promoter can be monitored.  
     
     
         17 . A vector according to  claim 16  wherein said proximal promoter region is linked to the coding region of the growth hormone gene (GH1).  
     
     
         18 . A vector according to  claim 16  wherein said proximal promoter region in said gene is further linked to a tag whereby the expression of said gene, and so the activity of said proximal promoter region, can be monitored.  
     
     
         19 . A vector according to  claim 18  wherein said tag is a protein tag.  
     
     
         20 . A vector according to  claim 10  which is further provided with at least one further proximal promoter region of the growth hormone gene (GH1).  
     
     
         21 . A vector according to  claim 20  wherein said additional proximal promoter region differs from that of the original proximal promoter region.  
     
     
         22 . A vector according to  claim 21  wherein each proximal promoter region is linked to a different coding sequence.  
     
     
         23 . A vector according to  claim 21  wherein each proximal promoter region is linked, either directly or indirectly, to a different tag that is capable of monitoring the activities of each of the said promoters.  
     
     
         24 . A host cell transformed with a vector according to  claim 10 .  
     
     
         25 . A host cell transformed with a vector according to  claim 15 .  
     
     
         26 . A host cell transformed with a vector according to  claim 16 .  
     
     
         27 . A host cell transformed with a vector according to  claim 20 .  
     
     
         28 . A recombinant cell line that is engineered to express a reporter molecule whose expression is under the control of the proximal promoter of the growth hormone gene wherein said proximal promoter comprises a plurality of the following SNP's: 1, 6, 7, 9, 11 or 14 and/or any one or more of the following haplotypes: 3, 4, 5, 7, 11, 13, 17, 19, 23, 24, 26 or 29 shown in Table 1.  
     
     
         29 . A transgenic non-human animal which under expresses growth hormone as a result of having a GH1 promoter containing a plurality of the following SNP's: 1, 6, 7, 9, 11 and 14 and/or as a result of said promoter being characterised by one of the following haplotypes: 3, 4, 5, 7, 11, 13, 17, 19, 23, 24, 26 or 29, shown in Table 1.  
     
     
         30 . A transgenic non-human animal according to  claim 29  wherein said promoter is characterised by haplotype 23.  
     
     
         31 . A transgenic non-human animal wherein the animal expresses growth hormone as a result of having a GH1 promoter characterised by haplotype 27.  
     
     
         32 . A transgenic non-human animal wherein the animal expresses growth hormone as a result of having a GH1 promoter characterised by haplotype 1.  
     
     
         33 . An artificial proximal promoter region of the growth hormone gene (GH1) characterised by the haplotype AGGGGTTAT-ATGGAG.  
     
     
         34 . An artificial proximal promoter region of the growth hormone gene (GH1) characterised by the haplotype AG-TTGTGGGACCACT.  
     
     
         35 . An artificial proximal promoter region of the growth hormone gene (GH1) characterised by the haplotype AG-TTTTGGGGCCACT.  
     
     
         36 . A method for screening therapeutically active drugs which can be used to treat growth hormone dysfunction comprising exposing a cell or cell line according to  claim 24 , to a candidate drug and then determining if the candidate drug has affected the activity of the promoter region of the growth hormone gene and so, in the case of the cell line, the expression of the reporter molecule.  
     
     
         37 . A method for screening therapeutically active drugs which can be used to treat growth hormone dysfunction comprising exposing a cell or cell line according to  claim 28 , to a candidate drug and then determining if the candidate drug has affected the activity of the promoter region of the growth hormone gene and so, in the case of the cell line, the expression of the reporter molecule.  
     
     
         38 . A method for screening for therapeutically active drugs which can be used to treat growth hormone dysfunction comprising exposing a transgenic non-human animal of  claim 29 , to candidate drugs and then monitoring the growth of said animal and where the candidate drug is shown to have a positive effect, in terms of animal growth, concluding that said growth is indicative of the therapeutic activity of said candidate drug.